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J P Dadoune

Publications and source records attributed to J P Dadoune.

At least 19 recordsLinked to original sources
PubMed2004

Identification of transcripts by macroarrays, RT-PCR and in situ hybridization in human ejaculate spermatozoa.

Round spermatids contain high levels of extremely varied mRNAs that are synthesized either throughout early spermatogenesis or during spermiogenesis from the haploid genome. Concomitantly, with major changes in the chromatin organization, arrest of transcription occurs at midspermiogenesis. However, previous investigations using RT-PCR have revealed the persistence of numerous and different transcripts in ejaculated spermatozoa. In the present study, a step-by-step analysis by means of macroarray hybridization, RT-PCR and in situ hybridization was performed to identify more accurately the different mRNA species found in the human ejaculated spermatozoa. The data showed an extended pattern of various transcripts encoding a diverse range of proteins involved in signal transduction and cell proliferation. For the first time, they demonstrated that mRNAs coding for the transcription factors NFkappaB, HOX2A, ICSBP, protein kinase JNK2, growth factor HBEGF and receptors RXRbeta and ErbB3 accumulate within the sperm nucleus. The origin and fate of the sperm transcripts remain subject to discussion.

Ejaculation↗
PubMed2001

Chromosomal factors of infertility in candidate couples for ICSI: an equal risk of constitutional aberrations in women and men.

To assess the frequency of chromosomal aberrations in French candidates for intracytoplasmic sperm injection (ICSI), and to explore the existence of a female chromosomal factor in some cases of couple infertility, a collaborative retrospective clinical and cytogenetic study was performed, launched by the Association des Cytogénéticiens de Langue Franciaise (ACLF). The karyotypes of 3208 patients [2196 men (68.4%), 1012 (31.6%) women] included in ICSI programmes over a 3-year period in France were collected. A total of 183 aberrant karyotypes was diagnosed, corresponding to an abnormality frequency of 6.1% (134/2196) for men and 4.84% (49/1012) for women. The following frequencies of abnormalities were observed respectively for men and women: 1.23% (n = 27) and 0.69% (n = 7) for reciprocal translocations, 0.82% (n = 18) and 0.69% (n = 7) for Robertsonian translocations, 0.13% (n = 3) and 0.69% (n = 7) for inversions, 3.32% (n = 73) and 2.77% (n = 28) for numerical sex chromosome aberrations, and 0.59% (n = 13) and 0% for other structural aberrations. Among the male patients of this latter group, 0.40% (n = 9) had a Y chromosome abnormality. Among the male patients with numerical sex chromosome abnormalities, 2.23% (n = 49) were 47,XXY, 0.32% (n = 7) were 47,XYY, and 0.77% (n = 17) had a mosaicism for numerical sex chromosome anomalies. All the female patients with sex chromosome abnormalities (2.77%, n = 28) had mosaicism for numerical sex chromosome anomalies. Even if these cases-the significance of which was sometimes questioned-were disregarded in the analysis, 2.08% (21/1012) of abnormal karyotypes remained in women. An overall increased frequency of chromosomal aberrations was found, and this confirmed that in some cases of poor reproductive outcome there may be a contribution of maternal chromosome aberrations. Indeed, the existence of a chromosome abnormality in the female partner was associated with the group of infertile men in which there was no apparent cause of infertility.

Chromosome Aberrations↗
PubMed2001

Expression of the TAR RNA binding protein in human testis.

In testis, several RNA binding proteins have been shown to play a role in the translational regulation of specific transcripts. The human protein TRBP (TAR RNA binding protein) is the homologue of the mouse Prbp (Prm-1 RNA binding protein) involved in the protamine mRNA translational delay. TRBP is known to activate the HIV-1 long terminal repeat but this protein has never been investigated during spermatogenesis. The aim of this work was to analyse the TRBP expression in human testis. By Northern blot analysis, we demonstrated a major 1.5 kb transcript present at a high level in human testis and, to a lesser extent, in some other tissues. In-situ hybridization revealed that this transcript was present only in elongating spermatids. Antibodies raised against a 27 amino acid TRBP-specific peptide revealed a single protein of 43 kDa expressed in the cytoplasm of elongated spermatids. At the ultrastructural level, quantitative analysis of both TRBP mRNA and protein, using electron microscopy in-situ hybridization and immunocytochemistry, showed that TRBP is expressed mainly in spermatids at steps 3-4 of spermiogenesis. These results are in agreement with the probable role of TRBP in the control of human protamine mRNA translation.

Gene Expression↗
PubMed2001

Accumulation of transcripts in the mature human sperm nucleus: implication of the haploid genome in a functional role.

The existence of sperm-specific transcripts has been suggested by a number of studies performed both in man and rodents but their origin and role are not yet elucidated. For evaluating the functional significance of these mRNAs, transcripts coding for proteins expressed during spermiogenesis or potentially implicated in the early steps of zygote development, have been searched in human testis and sperm cells by RT-PCR. Furthermore their localization in spermatozoa has been checked by in situ hybridization. Our results confirm the presence of basic nucleoproteins (Transition proteins 1 and 2, Protamines 1 and 2) spermatozoal transcripts which probably represent remnants of previous transcription. They also reveal the existence of sperm specific mRNAs coding for the transcription factor Stat 4, the cyclin B1 and for the testicular isozyme of the angiotensin converting enzyme ACE. On the contrary, mRNAs coding for the heat shock protein Hsp 70 have been found in testis but not in spermatozoa. The possible roles of these transcripts either during the fertilization process or in zygote are discussed.

Aged↗
PubMed2000

Sex chromosome mosaicism in males carrying Y chromosome long arm deletions.

Microdeletions of the long arm of the Y chromosome (Yq) are a common cause of male infertility. Since large structural rearrangements of the Y chromosome are commonly associated with a 45,XO/46,XY chromosomal mosaicism, we studied whether submicroscopic Yq deletions could also be associated with the development of 45,XO cell lines. We studied blood samples from 14 infertile men carrying a Yq microdeletion as revealed by polymerase chain reaction (PCR). Patients were divided into two groups: group 1 (n = 6), in which karyotype analysis demonstrated a 45,X/46,XY mosaicism, and group 2 (n = 8) with apparently a normal 46,XY karyotype. 45,XO cells were identified by fluorescence in-situ hybridization (FISH) using X and Y centromeric probes. Lymphocytes from 11 fertile men were studied as controls. In addition, sperm cells were studied in three oligozoospermic patients in group 2. Our results showed that large and submicroscopic Yq deletions were associated with significantly increased percentages of 45,XO cells in lymphocytes and of sperm cells nullisomic for gonosomes, especially for the Y chromosome. Moreover, two isodicentric Y chromosomes, classified as normal by cytogenetic methods, were detected. Therefore, Yq microdeletions may be associated with Y chromosomal instability leading to the formation of 45,XO cell lines.

Adult↗
PubMed2000

Y chromosome microdeletions and germinal mosaicism in infertile males.

Molecular deletions of the Y chromosome long arm are a frequent cause of male infertility. Because these deletions are thought to be inherited from fathers without Y chromosome deletions, the question arises as to whether their relatively high incidence in the male population could be due to the existence of a mosaicism in somatic and/or germinal paternal cells. This study included a total of 181 infertile men, among whom 18 were found to have an abnormal karyotype. In the other 163, polymerase chain reaction (PCR) analysis detected nine (5.5%) Y chromosome microdeletions. Blood, spermatozoa or testicular cells from 47 men (27 oligozoospermia, 20 azoospermia), including six Y-deleted patients, were screened for mosaicism using double target fluorescence in-situ hybridization (FISH) with Y centromeric and deleted in azoospermia (DAZ) gene-specific probes. Results indicated that: (i) percentages of double (intact Y chromosome) or single (deleted Y chromosome) fluorescent signals by FISH were in agreement with PCR data, thus demonstrating the reliability of the method; and (ii) a weak germ cell mosaicism was found in only two oligozoospermic patients, carrying 1.97 and 4.13% respectively of spermatozoa with a deleted Y chromosome. Further studies on larger populations are needed to evaluate precisely the incidence of Y deletion mosaicisms in infertile men.

Chromosome Aberrations↗
PubMed1999

Co-localization of HP1 and TP1 transcripts in human spermatids by double electron microscopy in situ hybridization.

Nuclear changes in the basic nucleoprotein complement occur during spermiogenesis in man. Somatic type histones are displaced by transition proteins which are replaced themselves by protamines, the major nuclear proteins present in late spermatids and sperm nuclei. Sense and antisense 35S-labelled riboprobes, coding respectively for human transition protein 1 (TP1) and protamine 1 (HP1), were synthesized with modified specific oligonucleotides and were used for light microscopy in situ hybridization. A double EM in situ hybridization was performed using a digoxigenin-labelled probe for TP1 and a biotin-labelled probe for HP1, and hybrids were revealed, respectively, with specific antibodies coupled to colloidal gold particles of different sizes (10 nm and 15 nm). For both types of transcripts, histological study revealed a specific distribution of the silver grains in the adluminal region of the seminiferous tubules where spermatids are localized. Quantitative ultrastructural analysis of the nuclear and cytoplasmic labelling densities for the mRNAs coding for TP1 and HP1 showed that the transcripts were found in both the nucleus and cytoplasm of round spermatids and persisted until the elongation phase. Transcripts accumulated in the spermatid cytoplasm without any particular cellular compartmentalization. At the end of the spermatid elongation phase, the disappearance of TP1 and HP1 transcripts may be related to the arrest of transcriptional activity, while the deposition of transition proteins and protamines occurs successively within spermatid nuclei.

Aged↗
PubMed1999

A high frequency of Y chromosome deletions in males with nonidiopathic infertility.

Microdeletions of the long arm of the human Y chromosome are associated with spermatogenic failure and have been used to define three regions of Yq (AZFa, AZFb, and AZFc) that are recurrently deleted in infertile males. In a blind study we screened 131 infertile males (46 idiopathic and 85 nonidiopathic) for Y chromosome microdeletions. Nineteen percent of idiopathic males, with an apparently normal 46,XY chromosome complement had microdeletions of either the AZFa, AZFb, or AZFc region. There was no strict correlation between the extent or location of the deletion and the phenotype. The AZFb deletions did not include the active RBM gene. Significantly, a high frequency of microdeletions (7%) was found in patients with known causes of infertility and a 46,XY chromosome complement. These included deletions of the AZFb and AZFc regions, with no significant difference in the location or extent of the deletion compared with the former group. It is recommended that all males with reduced or absence sperm counts seeking assisted reproductive technologies be screened for deletions of the Y chromosome.

Adult↗
PubMed1998

Advanced paternal age and de-novo complex chromosomal rearrangement in offspring.

We report one case of a de-novo complex chromosomal rearrangement (CCR), t(1;5;13)ins(14;13), in an abnormal 19-month-old boy. Clinical features associated were a mild facial dysmorphy and a psychomotor retardation. Parental ages were, respectively, 29 years for the mother and 60 years for the father. We point out the usefulness of fluorescence in-situ hybridization in elucidating CCRs, and discuss the possible correlation between the existence of a chromosomal aberration and advanced paternal age.

Adult↗
PubMed1998

Electron microscopic in situ hybridization study of simultaneous expression of TNP1 and PRM1 genes in human spermatids.

Nuclear changes in the basic nucleoprotein complement occur during human spermiogenesis. Somatic type histones are displaced by transition proteins which are replaced themselves by protamines, the major nuclear proteins found in late spermatids and spermatozoa nuclei. Digoxigenin or Biotin labeled probes, coding respectively for human transition protein 1 (TP1) and protamine 1 (HP1), were used for double EM in situ hybridization. Immunodetection of hybrids with specific antibodies coupled to colloidal gold particles of different size (10 nm and 15 nm) was performed on the same preparations. Quantitative analysis of the nuclear and cytoplasmic labeling densities for the mRNAs coding for TP1 and HP1 showed the presence of transcripts in both the nucleus and cytoplasm of round spermatids until the elongation phase. Transcripts accumulated in the spermatid cytoplasm without any particular cellular compartmentalization. At the end of the spermatid elongation phase, the disappearing of TP1 and HP1 transcripts may be related to the arrest of transcriptional activity while the deposition of transition proteins and protamines successively occurs within spermatid nuclei.

Aged↗
PubMed1997

Assisted reproductive technology and complex chromosomal rearrangements: the limits of ICSI.

Complex chromosomal rearrangements are very rare events in the human population. According to our knowledge on the consequences of simple reciprocal translocations for male fertility, translocations involving three or more chromosomes are thought to lead to severe reproductive impairments in terms of meiotic disturbance or chromosomal imbalance of gametes. We report the case of a 48 year old man whose sperm count revealed either oligozoospermia (<10(3) spermatozoa/ml) or azoospermia. He was referred to the laboratory for in-vitro fertilization after intracytoplasmic sperm injection. Cytogenetic investigations showed a complex chromosomal rearrangement involving firstly a translocation between the short arm of chromosome 7 and the long arm of chromosome 13 and secondly a translocation between the short arm of the same chromosome 13 and the short arm of chromosome 9. Diagnosis was ascertained by fluorescence in-situ hybridization and staining of the nucleolar organizer regions. Theoretical study of the translocated chromosomes predicted a 'chain' configuration of the hexavalent at the pachytene stage of meiosis. In all, 32 modes of segregation were considered and only one resulted either in a normal or a balanced gamete karyotype. Genetic counselling and choice of appropriate artificial reproduction technique are discussed.

Chromosomes, Human, Pair 13↗
PubMed1996

Immunoelectron microscopical distribution of histones H2B and H3 and protamines during human spermiogenesis.

The fine structural distribution of histones H2B and H3, and protamines were localized by means of specific antibodies and ultrastructural immunocytochemistry in nuclei of human spermatids and spermatozoa. The antibodies were used to detect the nuclear basic proteins on section of testis and ejaculated spermatozoa by immunoelectron microscopy. A quantitative analysis of labelling density was performed on micrographs using an interactive image analysis system. The labelling density of somatic-type histones H2B and H3 and of their testis-specific variants was constant in the nuclei of young spermatids with round nuclei (stages 1-2), and then increased in intermediate spermatids (stages 3-4). Histone H3 labelling decreased at the end of the elongation phase (stage 5) while histone H2B labelling decreased in mature spermatids (stage 6) only. Spermatozoa were found to be weakly labelled by the anti-histone antibodies. The first signs of labelling of protamines and basic intermediate proteins appeared in spermatid nuclei at stage 4, increased further in stage 6 spermatids and persisted in all sperm nuclei. The present work shows that histone-to-protamine replacement occurs at the beginning of the spermatid maturation phase in human. However, histones are partially retained in mature spermatids and sperm nuclei.

Adult↗
PubMed1995

A genetic study of angiotensin I-converting enzyme levels in human semen.

The plasma level of angiotensin I-converting enzyme (ACE) has been shown to be under genetic control. An insertion/deletion polymorphism in the ACE gene is associated with differences in the level of ACE in the plasma and inside T-lymphocytes. An ACE isoform is present in large amounts in spermatozoa and is expressed under an alternative, germ cell-specific promoter, whereas ACE present in the seminal fluid is the somatic form of ACE. We have investigated the effect associated with the I/D polymorphism on the level of ACE in seminal fluid and in spermatozoa. No differences in the level of ACE measured in the seminal fluid or in the spermatozoa were associated with the ACE I/D genotypes. We conclude that the modulation of expression associated with the I/D polymorphism is restricted to the somatic ACE promoter. These results also suggest that if one allele modulating the expression of ACE was under positive selection pressure, it was not through an effect on the semen concentration of ACE.

Alleles↗
PubMed1995

The nuclear status of human sperm cells.

In the last decade, and in particular since the development of in vitro fertilization techniques, the nuclear status of human sperm cells has shown to be a key parameter in the assessment of male fertility. The shape and condensed state of the mature sperm nucleus are determined by structural and functional events that occur during spermiogenesis. This paper reviews essential findings on re-organization of the nucleus during sperm differentiation and maturation, and reports recent data on the architecture, biochemical composition and stability of the nucleus in human ejaculated spermatozoa. Different methods used to evaluate nuclear maturity in relation to male fertility are critically appraised.

Cell Differentiation↗